Research Institute of Wildlife Ecology, Department for Interdisciplinary Life Sciences, Vetmeduni Vienna, Savoyenstraße 1, 1160, Vienna, Austria.
Konrad Lorenz Institute of Ethology, Department for Interdisciplinary Life Sciences, Vetmeduni Vienna, Savoyenstraße 1, 1160, Vienna, Austria.
Acta Parasitol. 2021 Jun;66(2):553-559. doi: 10.1007/s11686-020-00325-9. Epub 2020 Dec 21.
Alveolar echinococcosis is a severe helminthic disease in humans caused by larvae of the fox tapeworm Echinococcus multilocularis. Austria is considered an endemic area with hotspots having up to 45% prevalence (Bagó et al. in Proceedings of the Zoo and Wildlife Health Conference 2019, Berlin, p. 91, 2019). At our facility, we have registered a notifiable increase of animals submitted for the diagnosis of E. multilocularis since 2016. Therefore, we investigated high throughput diagnostic methods to provide rapid and reliable results in comparison with our current method.
We have developed and compared a novel method of detection using droplet digital PCR (ddPCR) combined with previous target specific extraction according to Maas et al. (Vet Parasitol 230:20-24, 2016), with our current macroscopic method "Shaking in a Vessel Technique" (SVT) by Duscher et al. (Parasitol Res 95(1):40-42, 2005). We investigated 77 wild canids (72 red foxes, 5 golden jackals) using both methods. The data were analyzed using a non-Bayesian approach, applying bootstrapping to create confidentiality intervals.
Sensitivity for droplet digital PCR was 90.51% with the 95% credibility interval ranging from 82.50 to 96.92%, whereas mean sensitivity for SVT was 92.04% with a 95% credibility interval ranging from 84.75% to 98.36%. Additionally, a non-linear regression similar to R could be pointed out between the counted worms and the results gathered from ddPCR.
Magnetic capture extraction followed by ddPCR shows strong potential as a high throughput method for diagnosing E. multilocularis prevalence in diverse canid populations as well as infection intensities of individual animals, giving valuable epidemiological insights of the distribution amongst wild canids as an alternative to conventional qPCR or macroscopic methods.
泡型包虫病是一种严重的人类寄生虫病，由狐绦虫多房棘球蚴的幼虫引起。奥地利被认为是一个流行地区，热点地区的患病率高达 45%（Bagó 等人在 2019 年柏林动物园和野生动物健康会议记录中，第 91 页，2019 年）。在我们的设施中，自 2016 年以来，我们已经记录到提交用于诊断多房棘球蚴的动物数量有显著增加。因此，我们研究了高通量诊断方法，以便与我们目前的方法相比提供快速可靠的结果。
我们开发并比较了一种新的检测方法，使用液滴数字 PCR（ddPCR）结合 Maas 等人之前提出的针对特定目标的提取方法（兽医寄生虫学 230:20-24, 2016），与我们目前的宏观方法“容器摇动技术”（SVT）由 Duscher 等人提出（寄生虫学研究 95(1):40-42, 2005）。我们使用这两种方法对 77 只野生犬科动物（72 只红狐，5 只金豺）进行了研究。使用非贝叶斯方法分析数据，通过自举法创建置信区间。
ddPCR 的灵敏度为 90.51%，95%置信区间范围为 82.50%至 96.92%，而 SVT 的平均灵敏度为 92.04%，95%置信区间范围为 84.75%至 98.36%。此外，可以指出，与 ddPCR 相比，计数的蠕虫数量与从 ddPCR 获得的结果之间存在类似于 R 的非线性回归。
磁捕获提取后进行 ddPCR 显示出作为一种高通量方法的强大潜力，可用于诊断不同犬科种群中的多房棘球蚴流行率以及单个动物的感染强度，为野生犬科动物的分布提供有价值的流行病学见解，作为传统 qPCR 或宏观方法的替代方法。
